Oestradiol-2-hydroxylase (E2-OH) activity was measured in liver and brain microsomes of 6-8-week-old Wistar rats. Phenobarbitone (75 mg/kg daily for 3 days) significantly increased enzyme activity in the liver of males and females, but there were striking differences between the two sexes. In males the enzyme activity was increased by 37% over control values and in females by 200%. The total microsomal cytochrome P-450 content was increased by 75% in males and by 82% in females. The apparent Michaelis constant (Km) of E2-OH for 17β-oestradiol in untreated males (9.8 μM) and females (9.2 μM) did not differ significantly. Phenobarbitone treatment, however, tended to reduce the apparent Km in males (8.2 μM) and to increase it in females (18.7 μM). E2-OH activity was also detected in brain tissue of both sexes, but it was 50-200-fold lower than in the liver and was not increased by phenobarbitone.